title: Transcriptional and post-transcriptional regulation by nickel of sodN gene encoding nickel-containing superoxide dismutase from Streptomyces coelicolor Müller.
abstract: A novel type of superoxide dismutase containing nickel as a cofactor (NiSOD) has been discovered in several Streptomyces spp. The gene for NiSOD (sodN) was cloned from S. coelicolor Müller using degenerate oligonucleotide probes designed from the N-terminal peptide sequence of the purified enzyme. It encodes a polypeptide of 131 amino acids (14703 Da), without any apparent sequence similarity to other known proteins. The N-terminus of the purified NiSOD was located 14 amino acids downstream from the initiation codon of the deduced open reading frame (ORF), indicating the involvement of protein processing. The molecular mass of the processed polypeptide was predicted to be 13201 Da, in close agreement with that of the purified NiSOD (13.4 kDa). The transcription start site of the sodN gene was determined by S1 mapping and primer extension analysis. Ni2+ regulates the synthesis of NiSOD polypeptide. S1 mapping of both 5' and 3' ends of sodN mRNA revealed that Ni2+ increased the level of monocistronic sodN mRNA by more than ninefold without changing its half-life, thus demonstrating that Ni2+ regulates transcription. Both precursor and processed NiSOD polypeptides with little SOD activity were produced from the cloned sodN gene in S. lividans in the absence of sufficient Ni2+; however, on addition of Ni2+, active NiSOD consisting of only processed polypeptide was formed. Expression of the full-length sodN gene in E. coli produced NiSOD polypeptide without any SOD activity even in the presence of Ni2+. However, deletion of nucleotides encoding the N-terminal 14 amino acids from the sodN gene allowed the production of active NiSOD in E. coli, indicating that N-terminal processing is required to produce active NiSOD. These results reveal the unique role of nickel as a multifaceted regulator in S. coelicolor controlling sodN transcription and protein processing, as well as acting as a catalytic cofactor.
authors.affiliation: Department of Microbiology, College of Natural Sciences, Seoul National University, Korea.
authors.forename: E J
authors.initials: EJ
authors.lastname: Kim
authors.forename: H J
authors.initials: HJ
authors.lastname: Chung
authors.forename: B
authors.initials: B
authors.lastname: Suh
authors.forename: Y C
authors.initials: YC
authors.lastname: Hah
authors.forename: J H
authors.initials: JH
authors.lastname: Roe
mesh_terms: D000595:Amino Acid Sequence; D000596:Amino Acids / Q000032:analysis; D001483:Base Sequence; D015153:Blotting, Western; D003001:Cloning, Molecular; D004587:Electrophoresis, Agar Gel; D004591:Electrophoresis, Polyacrylamide Gel; D015964:Gene Expression Regulation, Bacterial / Q000235:genetics*; D015971:Gene Expression Regulation, Enzymologic / Q000235:genetics*; D008969:Molecular Sequence Data; D009532:Nickel / Q000737:chemistry / Q000502:physiology*; D009693:Nucleic Acid Hybridization; D016366:Open Reading Frames / Q000235:genetics; D012323:RNA Processing, Post-Transcriptional; D012329:RNA, Bacterial / Q000737:chemistry / Q000378:metabolism; D012333:RNA, Messenger / Q000737:chemistry / Q000378:metabolism; D015719:Single-Strand Specific DNA and RNA Endonucleases / Q000737:chemistry / Q000494:pharmacology; D013302:Streptomyces / Q000201:enzymology* / Q000235:genetics; D013482:Superoxide Dismutase / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D014158:Transcription, Genetic
chemical_list: D000596:Amino Acids; D012329:RNA, Bacterial; D012333:RNA, Messenger; D009532:Nickel; D013482:Superoxide Dismutase; D015719:Single-Strand Specific DNA and RNA Endonucleases